Giachetti C, Linnen J M, Kolk D P, Dockter J, Gillotte-Taylor K, Park M, Ho-Sing-Loy M, McCormick M K, Mimms L T, McDonough S H
Research and Development, Gen-Probe Incorporated, San Diego, California 92121, USA.
J Clin Microbiol. 2002 Jul;40(7):2408-19. doi: 10.1128/JCM.40.7.2408-2419.2002.
Various nucleic acid assays have been developed and implemented for diagnostics and therapeutic monitoring of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) infections. The high-throughput, semiautomated assays described here were developed to provide a method suitable for screening plasma specimens for the presence of HIV-1 and HCV RNAs. Three assays were developed: a multiplex HIV-1/HCV assay for simultaneous detection of HIV-1 and HCV, and discriminatory assays for specific detection of HIV-1 and HCV. The assay systems utilize three proprietary technologies: (i) target capture-based sample preparation, (ii) transcription-mediated amplification (TMA), and (iii) hybridization protection assay (HPA). An internal control is incorporated into each reaction to control for every step of the assay and identify random false-negative reactions. The assays demonstrated a sensitivity of at least 100 copies/ml for each target, and they detected with similar sensitivity all major variants of HCV and HIV-1, including HIV-1 group O strains. Assay sensitivity for one virus was not affected by the presence of the other. The specificity of these TMA-driven assays was >or=99.5% in both normal donor specimens and plasma containing potentially interfering substances or other blood-borne pathogens. Statistical receiver operating characteristic plots of 1 - specificity versus sensitivity data determined very wide analyte cutoff values for each assay at the point at which the assay specificity and sensitivity were both >or=99.5%. The sensitivity, specificity, and throughput capability predict that these assays will be valuable for large-volume plasma screening, either in a blood bank setting or in other diagnostic applications.
已经开发并实施了各种核酸检测方法，用于人类免疫缺陷病毒1型（HIV-1）和丙型肝炎病毒（HCV）感染的诊断和治疗监测。本文所述的高通量、半自动检测方法旨在提供一种适用于筛查血浆标本中HIV-1和HCV RNA存在情况的方法。开发了三种检测方法：一种用于同时检测HIV-1和HCV的多重HIV-1/HCV检测方法，以及用于特异性检测HIV-1和HCV的鉴别检测方法。检测系统采用了三种专有技术：（i）基于靶标捕获的样品制备，（ii）转录介导的扩增（TMA），以及（iii）杂交保护分析（HPA）。每个反应中都加入了一个内部对照，以控制检测的每一步，并识别随机的假阴性反应。这些检测方法对每个靶标的灵敏度至少为100拷贝/毫升，并且以相似的灵敏度检测到HCV和HIV-1的所有主要变体，包括HIV-1 O组毒株。一种病毒的检测灵敏度不受另一种病毒存在的影响。在正常供体标本以及含有潜在干扰物质或其他血源性病原体的血浆中，这些由TMA驱动的检测方法的特异性均≥99.5%。在检测特异性和灵敏度均≥99.5%的点上，1-特异性与灵敏度数据的统计接收器操作特征图确定了每种检测方法非常宽的分析物临界值。灵敏度、特异性和通量能力表明，这些检测方法对于血库环境或其他诊断应用中的大量血浆筛查将具有重要价值。
